DETERMINATION OF ANTIGEN-SPECIFIC - MEMORY EFFECTOR CD4+ T-CELL FREQUENCIES BY FLOW-CYTOMETRY - EVIDENCE FOR A NOVEL, ANTIGEN-SPECIFIC HOMEOSTATIC MECHANISM IN HIV-ASSOCIATED IMMUNODEFICIENCY/

The highly regulated secretion of effector cytokines by CD4+ T cells p lays a critical role in immune protection against pathogens such as cy tomegalovirus. Here, we directly compare the frequency and functional characteristics of cytomegalovirus-specific CD4+ memory/effector T cel ls in normal and HIV+ subjects using a novel, highly efficient multipa rameter flow cytometric assay that detects the rapid intracellular acc umulation of cytokine(s) after short-term (6 h) in vitro antigen stimu lation. Responses in this assay correlate precisely with independent m easures of sensitization history (e.g., seroreactivity), and allow the simultaneous assessment of multiple cytokines in single effector T ce lls, Healthy HIV- individuals manifested an average of 0.71, 0.72, 0.3 8, and 0.06% CD4+ T cells responding to cytomegalovirus with gamma-IFN , TNF-alpha, IL-2, and IL-4 production, respectively, with the simulta neous production of gamma-IFN, TNF-alpha, and IL-2 being the most comm on effector phenotype. Significantly, overall cytomegalovirus-specific CD4+ effector frequencies were markedly higher among 40% of HIV+ subj ects (2.7-8.0%), and demonstrated a predominately polarized gamma-IFN/TNF-alpha+/IL-2-/IL-4- phenotype, In contrast, CD4+ effector frequenc ies for heterologous, nonubiquitous viruses such as the mumps virus we re low or absent in the HIV+ group. These data suggest the existence o f homeostatic mechanisms in HIV disease that selectively preserve memo ry T cell populations reactive with ubiquitous pathogens such as cytom egalovirus-likely at the expense of T cell memory to more sporadically encountered infectious agents.